In monitoring medical conditions and the response of patients to treatment efforts, it is desirable to use analytical methods that are fast, accurate, and convenient for the patient. Electrochemical methods have been useful for quantifying certain analytes in body fluids, particularly in blood samples. Typically, these biological analytes, such as glucose, undergo redox reactions when in contact with specific enzymes. The electric current generated by such redox reactions may be correlated with the concentration of the biological analyte in the sample.
Tiny electrochemical cells have been developed that provide patients the freedom to monitor blood analyte concentrations without the need of a healthcare provider or clinical technician. Typical patient-operated electrochemical systems utilize a disposable sensor strip with a dedicated measurement device containing the necessary circuitry and output systems. For analysis, the measurement device is connected to the disposable electrochemical sensor strip containing the electrodes and reagents to measure the analyte concentration in a sample that is applied to the strip.
The most common of these miniature electrochemical systems are glucose sensors that provide measurements of blood glucose levels. Ideally, a miniature sensor for glucose should provide accurate readings of blood glucose levels by analyzing a single drop of whole blood, typically from 1-15 microliters (μL).
In a typical analytical electrochemical cell, the oxidation or reduction half-cell reaction involving the analyte produces or consumes electrons, respectively. This electron flow can be measured, provided the electrons can interact with a working electrode that is in contact with the sample to be analyzed. The electrical circuit is completed through a counter electrode that is also in contact with the sample. A chemical reaction also occurs at the counter electrode, and this reaction is of the opposite type (oxidation or reduction) relative to the type of reaction at the working electrode. Thus, if oxidation occurs at the working electrode, reduction occurs at the counter electrode. See, for example, Fundamentals Of Analytical Chemistry, 4th Edition, D. A. Skoog and D. M. West; Philadelphia: Saunders College Publishing (1982), pp 304-341.
Some conventional miniaturized electrochemical systems include a true reference electrode. In these systems, the true reference electrode may be a third electrode that provides a non-variant reference potential to the system, in addition to the working and counter electrodes. While multiple reference electrode materials are known, a mixture of silver (Ag) and silver chloride (AgCl) is typical. The materials that provide the non-variant reference potential, such as a mixture of silver and silver chloride, are separated, by their insolubility or other means, from the reaction components of the analysis solution.
In other miniature electrochemical systems, a combination counter/reference electrode is employed. These electrochemical sensor strips are typically two electrode systems having a working electrode and a counter/reference electrode. The combined counter/reference electrode is possible when a true reference electrode also is used as the counter electrode.
Because they are true reference electrodes, counter/reference electrodes are typically mixtures of silver (Ag) and silver chloride (AgCl), which exhibit stable electrochemical properties due to the insolubility of the mixture in the aqueous environment of the analysis solution. Since the ratio of Ag to AgCl does not significantly change during transient use, the potential of the electrode is not significantly changed.
An electrochemical sensor strip is typically made by coating a reagent layer onto the conductor surface of an analysis strip. To facilitate manufacturing, the reagent layer may be coated as a single layer onto all of the electrodes.
The reagent layer may include an enzyme for facilitating the oxidation or reduction reaction of the analyte, as well as any mediators or other substances that assist in the transfer of electrons between the analyte reaction and the conductor surface. The reagent layer also may include a binder that holds the enzyme and mediator together, thus allowing them to be coated onto the electrodes.
Whole blood (WB) samples contain red blood cells (RBC) and plasma. The plasma is mostly water, but contains some proteins and glucose. Hematocrit is the volume of the RBC constituent in relation to the total volume of the WB sample and is often expressed as a percentage. Whole blood samples generally have hematocrit percentages ranging from 20 to 60%, with ˜40% being the average.
One of the drawbacks of conventional electrochemical sensor strips utilized to measure the glucose concentration in WB is referred to as the “hematocrit effect.” The hematocrit effect is caused by RBC blocking the diffusion of the mediator or other measurable species to the conductor surface for measurement. Because the measurement is taken for the same time period each time a sample is tested, blood samples having varying concentrations of RBC can cause inaccuracies in the measurement. This is true because the sensor cannot distinguish between a lower measurable species concentration and a higher measurable species concentration where the RBC interfere with diffusion of the measurable species to the conductor surface. Thus, variances in the concentration of RBC in the WB sample result in inaccuracies (the hematocrit effect) in the glucose reading.
If WB samples containing identical glucose levels, but having hematocrits of 20, 40, and 60%, are tested, three different glucose readings will be reported by a conventional system that is based on one set of calibration constants (slope and intercept, for instance). Even though the glucose concentrations are the same, the system will report that the 20% hematocrit sample contains more glucose than the 60% hematocrit sample due to the RBC interfering with diffusion of the measurable species to the conductor surface.
Conventional systems are generally configured to report glucose concentrations assuming a 40% hematocrit content for the WB sample, regardless of the actual hematocrit content in the blood sample. For these systems, any glucose measurement performed on a blood sample containing less or more than 40% hematocrit will include some inaccuracy attributable to the hematocrit effect.
Conventional methods of reducing the hematocrit effect for amperometric sensors include the use of filters, as disclosed in US. Pat. Nos. 5,708,247 and 5,951,836; reversing the potential of the read pulse, as disclosed in WO 01/57510; and by methods that maximize the inherent resistance of the sample, as disclosed in U.S. Pat. No. 5,628,890. While each of these methods balance various advantages and disadvantages, none are ideal.
As can be seen from the above description, there is an ongoing need for improved devices and methods for determining the concentration of biological analytes, including glucose. The devices and methods of the present invention may decrease the error introduced by the hematocrit and other effects in WB samples.